Ninety percent of those diagnosed with systemic lupus erythematosus (SLE) are women, with peak incidence between the ages of 15 and 45, when women are most hormonally active. Despite significant research effort, the mechanisms underlying this sex bias remain unclear. Our laboratory previously backcrossed estrogen receptor alpha knockout (ER?KO) mice onto the NZM2410 lupus prone background. We demonstrated that female NZM/ER?KO mice had significantly less renal disease and significantly prolonged survival compared to WT littermates despite similar serum autoantibodies and glomerular immune complex deposition. ER?KO mice are not ER? null, but rather express an N-terminally truncated ER?. They have physiologic deficiencies including infertility due to disruption of a critical activation domain (AF-1). We showed that dendritic cell (DCs) from NZM/ER?KO mice have a blunted inflammatory response to Toll-like receptor (TLR) ligands. When these mice were ovariectomized, the protective phenotype was lost. Upon estradiol-repletion, protection was restored. True ER? null mice are not protected, suggesting that estrogen in the presence of the AF-1 mutant confers protection, rather than the absence of the full-length ER?66. Interestingly, the truncated ER? expressed in the ER?KO animal is structurally similar to ER?46, an endogenous ER? splice variant that lacks the AF-1 domain, and is a negative regulator of gene transcription. ER?46 has an identical DNA binding domain to ER?66 and is a powerful inhibitor of ER?66. We hypothesize that ER?46 expression has a protective effect in lupus. The goal of this project is to determine the role of ER?46 in SLE and TLR-induced inflammation and to improve our understanding of ER?-mediated transcription in the setting of inflammation. In order to accomplish this, Dr. Cunningham will (1) Define the in vitro molecular mechanisms underlying ER?46- and ER?66-regulated transcriptional activity impacting the innate inflammatory response of murine DCs, (2) Determine ex vivo and in vivo the role of ER?46 in TLR-induced lupus utilizing a murine strain expressing an A/B truncation mutant of ER? (ER?AF10), and (3) Use human peripheral blood monocyte (PBMC)-derived DCs to define the role of ER?46 and ER?66 in regulation of the innate immune response in DCs from controls vs. lupus patients. Dr. Cunningham is a clinician-investigator with a long-term career goal of becoming an independent basic and translational researcher in the field of immunology with a focus on SLE. To facilitate her transition to independence, she seeks to further her training in dendritic cell biology, ChIPseq and bioinformatics, and the use of human cells from patient samples. Dr. Cunningham has a mentorship team with an outstanding mentoring track record and wide expertise to support her project. The studies proposed in this K08 mentored grant application will significantly advance the field by providing understanding of ER?-mediated transcription in the setting of TLR-induced inflammation, including ER? binding partners, gene targets, and modulation by ER?46, which represents a potential therapeutic agent.